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张 超,吴俊学,王 冶,谢海洋,彭 玲.和厚朴酚对 D-半乳糖致腰椎间盘退变大鼠血清炎症因子及细胞线粒体凋亡通路的影响[J].中国比较医学杂志,2021,31(11):62~68,75.
和厚朴酚对 D-半乳糖致腰椎间盘退变大鼠血清炎症因子及细胞线粒体凋亡通路的影响
Effects of honokiol on serum inflammatory factors and the mitochondrial apoptosis pathway in rats with D-galactose-induced intervertebral disc degeneration
投稿时间:2020-11-10  
DOI:10. 3969 / j.issn.1671-7856. 2021. 11. 010
中文关键词:  和厚朴酚  腰椎间盘退变  炎症因子  凋亡
英文关键词:honokiol  intervertebral disc degeneration  inflammatory factor  apoptosis
基金项目:
作者单位E-mail
张 超 川北医学院附属医院 骨外科,四川 南充 637000 zhangchaokaituo@ 163.com 
吴俊学 川北医学院附属医院 骨外科,四川 南充 637000  
王 冶 川北医学院附属医院 骨外科,四川 南充 637000  
谢海洋 川北医学院附属医院 骨外科,四川 南充 637000  
彭 玲 川北医学院附属医院 临床营养科,四川 南充 637000 pengling199001@ 163.com 
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中文摘要:
       目的 探索和厚朴酚对 D-半乳糖致腰间盘退变大鼠血清炎症因子含量变化,细胞外基质标记物表达水平及凋亡通路相关蛋白表达水平的影响。 方法 45 只 SD 大鼠随机分为对照组( Control)、D-半乳糖(D- galactose,D-gal)模型组(Model)、和厚朴酚低剂量组(2 mg / kg)、和厚朴酚中剂量组(4 mg / kg)、和厚朴酚高剂量组 (8 mg / kg),每组 9 只。模型组和和厚朴酚给药组采用皮下注射 D-gal方法建立大鼠椎间盘退变模型,对照组皮下注射等体积生理盐水。 造模成功后,给药组大鼠分别灌胃和厚朴酚 2、4、8 mg / kg,其余各组大鼠灌胃生理盐水,连续给药 4 周后处死大鼠。 HE 和番红 O 染色观察比较各组腰椎间盘形态组织学;qRT-PCR 检测大鼠椎间盘组织细胞外基质标记物的 mRNA 水平;ELISA 检测大鼠血清中白细胞介素 IL-1β、IL-6、IL-10 和肿瘤坏死因子 α(TNF-α)含量的变化;TUNEL 染色观察大鼠椎间盘的组织凋亡情况;Western blot 检测 B 细胞淋巴瘤-2(Bcl-2) / Bcl-2 相关 X 蛋白(Bax)、胱天蛋白酶-9(Caspase-9)、Caspase-3、c-Myc 的蛋白表达。 结果 模型组和和厚朴酚低、中剂量组椎间盘组织形态学评分较对照组显著升高(P<0. 05)。 和厚朴酚中、高剂量组椎间盘组织形态学评分较模型组显著降低 (P<0. 05)。 模型组和和厚朴酚低剂量组软骨受损严重,和厚朴酚中、高剂量组软骨受损较轻。和厚朴酚能显著升高 SRY 相关 HMG 盒 9(SOX-9)、II 型胶原(Collagen II)和蛋白聚糖(Aggrecan)的表达水平(P<0. 05)。模型组和和厚朴酚低、中、高剂量组 IL-1β、IL-6、IL-10 和 TNF-α 的含量较对照组均显著升高(P<0. 05)。与模型组比较,和厚朴酚中、高剂量组 IL-1β、IL-6 和 TNF-α 含量均显著降低(P<0. 05);IL-10 的含量显著升高(P<0. 05)。 与对照组相比, 模型组和和厚朴酚低、中剂量组凋亡细胞数显著升高(P<0. 05),Bcl-2/ Bax 比值显著降低(P<0. 05),Caspase-9、 Caspase-3、c-Myc 的蛋白水平均显著升高(P<0. 05)。 与模型组相比,和厚朴酚中、高剂量组凋亡细胞数显著降低(P<0. 05),Bcl-2/ Bax 比值显著升高(P<0. 05),Caspase-9、Caspase-3、c-Myc 的蛋白水平均显著降低(P<0. 05)。 结论 和厚朴酚对 D-半乳糖致腰椎间盘退变有明显疗效。
英文摘要:
       Objective To explore the effect of honokiol on serum inflammatory factors, extracellular matrix component expression, and apoptotic pathway-related protein expression in rats with D-galactose-induced intervertebral disc degeneration. Methods Forty-five SD rats were randomly divided into control ( Control), D-galactose ( D-gal) model (Model), honokiol low dose (2 mg / kg), honokiol medium dose (4 mg / kg) and honokiol high dose (8 mg / kg) groups, with 9 rats in each group. The rat model of intervertebral disc degeneration was established by subcutaneous injection of D- galactose in model and honokiol groups, and the Control group was subjected to subcutaneous injection of normal saline. After successful modeling, administration group rats were gavaged with magnolol at 2, 4 and 8 mg / kg, and the remaining rats were gavaged with normal saline in each group. Rats were sacrificed after 4 weeks of continuous administration. HE and safranin O staining was used to observe and compare the morphology and histology of intervertebral discs in each group. The mRNA levels of extracellular matrix components were measured by qRT-PCR in rat intervertebral discs. Changes of interleukin IL-1β, IL-6, tumor necrosis factor-α ( TNF-α) and IL-10 in rat serum were detected by ELISAs. TUNEL staining was performed to observe apoptosis of rat intervertebral disc. Protein expression of B-cell lymphoma protein 2(Bcl- 2) / Bcl-2 associated X protein ( Bax), Caspase-9, Caspase-3 and c-Myc was detected by Western blot. Results The scores of intervertebral disc histomorphology in the D-galactose group and honokiol low and medium dose groups were significantly higher than that in the control group (P<0.05). Compared with that in the D-galactose group, the scores of intervertebral disc histomorphology in medium and high dose honokiol groups were significantly lower (P<0.05). Cartilage damage in the model group and the honokiol low dose group was severe, and that in honokiol medium and high dose groups was slightly damaged. Honokiol significantly increased the expression levels of SRY-related high mobility group-box gene 9 ( SOX-9), collagen II and aggrecan (P<0.05). The levels of IL-1β, IL-6, TNF-α and IL-10 in the D-galactose group and honokiol medium and high dose groups were significantly higher than those in the control group (P<0.05). Compared with that in the D-galactose group, the levels of IL-1β, IL-6 and TNF-α in the honokiol medium and high dose groups were reduced significantly (P<0.05), and the content of IL-10 was increased significantly (P<0. 05). Compared with that in the control group, the number of apoptotic cells in the model group and honokiol low and medium dose groups was increased significantly (P< 0. 05), the Bcl-2/ Bax ratio decreased significantly (P< 0.05), and the protein levels of Caspase-9, Caspase-3 and c-Myc were increased significantly ( P< 0.05). Compared with that in the model group, the number of apoptotic cells in the honokiol medium and high dose groups was decreased significantly (P<0. 05), the Bcl-2/ Bax ratio was increased significantly (P<0.05), and the protein levels of Caspase-9, Caspase-3 and c-Myc were decreased significantly (P<0.05). Conclusions Honokiol has obvious therapeutic effects on lumbar intervertebral disc degeneration induced by D-galactose.
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